| Objective: Ache is the commom symptom of many diseases in clinic. At present, most of the analgesics used in clinic therapy had addiction and obvious adverse drug reaction, so it is far-reaching significant to develop a high-effect and low-toxicity analgesic. Yuanhu Zhitong Pian composed by CORYDALIS YANHUSUO W. T. WANG and ANGELICA DAHURICA is recorded in Chinese Pharmacopeia since 1985 for its simple anagraph and difinite effect, it has obvious analgesic effect without the common toxicity and adverse drug reaction of chemicals, especially absence of dependence. At present, CORYDALIS YANHUSUO W. T. WANG and ANGELICA DAHURICA were extracted by alcohol refluxing method in pharmaceutical industry, which exists many limitations such as mass extract, consuming lots of solvent etc. Available Yuanhu Zhitong preparations are inadvanced in preparation form and absent in quality control, so it is necessary to improve extraction, preparation form and quality control of available YuanhuZhitong preparation. Methods:The CO2-SFE of CORYDALIS YANHUSUO W. T. WANG (1)The content of tetrahydropalmatine(THP) was determined by HPLC, which was considered as a quality control of extraction process; (2)To compare the THP extraction rate of different alkalinized methods; (3)On the basis of mono-factor testing, uniform design was used to optimize the extraction technique of CORYDALIS YANHUSUO W. T. WANG , which was evaluated by comparing the content of THP; (4)The comparison was done between CO2-SFE and alcohol refluxing method for the extraction effect of effective components from CORYDALIS YANHUSUO W. T. WANG.The COrSFE of ANGELICA DAHURICA ?The contents of imperator(IMP) and isoimperatorin(iso-IMP) were determined by HPLC, which was considered as the quality control of extraction process; ?On the basis of mono-factor testing, orthogonal design was used to investigate the impact factors on CO2-SFE; ?The comparison was done between CO2-SFE method and alcohol refluxing method for the extraction effect of effective components from ANGELICA DAHURICA.Preparation of dispersible tablets d)The accessories were selected through mono-factor testing by observing tablet formation, dispersible uniformity and disintegrity time; (D Uniform design was exploited to optimize the preparation technique by determining the disintegrity time and TJ0 of THP; ?The reproducibility of preparing process was evaluated by the contents determination and dispersible uniformity test. The quality standards of dispersible tablet was established.Stability test of dispersible tablets ?To find the factors affecting stability by observing the change of out-look of the dispersible tablet and determining disintegrity time, contents of THP and IMP; ?In the accelerated testing, the expiratory time was estimated by observing the change of appearance and determining T50 of THP, the contents of THP and IMP.The dispersible tablets and conventional tablets was compared by dissolution experiment in vitro, pharmacokinetics experiment in dogs and pharmacodynamics in mice.Results:The COrSFE of CORYDALIS YANHUSUO W. T. WANG ?There was a good linear relation for THP at the range of 9 u g/ml72 v- g/ml(r=0.9999), the precise RSD of this method was less than 3.0%,the mean recovery was 102.41%. Method exploited can serve to determine the content of THP in the CO2-SFE extract from CORYDALIS YANHUSUO W. T. WANG. ?The extracted rate of THP was the highest in the sample alkalinized with saturation Ca(OH)2 solution before CO2-SFE; ?The optimum technique paramete for extracting alkaloid from CORYDALIS YANHUSUO W. T. WANG were 40 mesh particle, 45MPa as extraction pressure, 60*C as extraction tempreture, 1.4h as the extraction time with a modifier of anhydrousalcohol; ?The target components extracted by CO2-SFE and alcohol refluxing method from CORYDALIS YANHUSUO W. T. WANG were the same. The CO2-SFE extract was more purer.the content of THP was 2.3 times and the weight of solid after dehydration was one fourth of the amount obtained by traditional method.The CO2-SFE of ANGELICA DAHURICA ?The linear relation was good for IMP at the range of 29.4—235.2 u g/ml (r=0.9999) and for iso-IMP at the range of 10—80 w g/ml (r=0.9999). The precise RSD was less than 2.0% for either IMP or iso-IMP, the mean recovery was 100.46% for IMP and 99.94% for iso-IMP.The established method can serve to determine the contents of IMP and iso-IMP in the CO2-SFE extract from ANGELICA DAHURICA. ?The extraction rate of total coumarins from ANGELICA DAHURICA was highest when the sample was pulverized into 60 mesh and extracted at 35MPa and 45*C for 3h with a modifier of anhydrous alcohol. For ANGELICA DAHURICA, the extraction pressure and pulverized degree can affect on the extraction very remarkably and the extraction time can do remarkably. (3)The target components from ANGELICA DAHURICA extracted by CO2-SFE and alcohol refluxing method were the same.The CO2-SFE extract was more purer,the content of total coumarins was a little higher than that of traditional method, and the weight of solid after dehydration was one sixth of that extracted by the traditional method.Preparation of dispersible tablets ?The weight of dispersible tablet was definited as 400mg, the accessories were 34% starch, 40%MCC and 10%lactose as filler, cCMC-Na as disintegration, PVPK30 alcohol solution as adhesive agents on the basis of mono-factor test. ?The optimal concentration was 8% for cCMC-Na and was 15% for PVPK30 alcohol solution selected through uniform design. ?The average contents of THP and imperitor of three batches are 3.49mg/g and 2.81mg/g, respectively. The raw herb medicine contained in each dispersible tablet are correspond to that in five conventional tablets. The dispersible uniformity of the three batches accorded with demand of Chinese pharmacopeia(2000 edition). The items of quality standards of dispersible tablet include appearance, CO2-SFE process of two TCM, preparation, identification, examination, content determination, etc.Stress test showed that high tempreture, high humidity and lightening can decrease the content of THP and high humidity can decrease the content of IMP. The content of THP decreased more than 10% after 2 monthes and that of IMP decreased more than 10% after 3 monthes in the accelerated test. Yuanhu Zhitong dispersible tablet should be packed in a sealed and light-resistant package and stored in a cool place.Comparison of the dispersible tablets and conventional tablets ?The dissolution rate of dispersible tablet was affected greatly by pH. THP can dissolved completely in solution pHl, which was taken as dissolving media; ?The dissolving speed of THP from dispersible tablet was fast and more than 85% of THP was dissolved in 12 minutes, but the THP in conventional tablet began to dissolved at 20 minutes and less than 80% of THP was dissolved in 60 minutes; ?The dissolution curve of THP in dispersible tablets in vitro can be well imitated by first-order model or Weibull distributing model, and that of conventional tablet by zero-order model; ? The plasm data were processed by DAS verl.O (Drug And Statistics for Windows), THP in vivo was described by one-component model. The paramete of dispersible tablet in dogs as following: lm 1.23h, Tmax 0.75h, Cmax 269.8 u g/L, AUC 519.85 u g/(Lh). The concentration of THP in conventional tablet in vivo is undetectable in most time, so the paramete of conventional tablet can't be calculated. From plasm drug concentration-time curve, differency can be observed that THP absorption of dispersible tablet was fast obviously, Tmax was faster, Cmax was higher and AUC was larger than those of conventional tablet, there was a remarkable difference between the dispersible and conventional tablets. ?In hot plate test, there was a very remarkable difference between high dosage of dispersible tablet and the same dosage of conventional tablet(P<0.01 M 0.001),and remarkable difference between low dosage of dispersible tablet and that of conventional tablet (P<0.01 s￡ 0.001);in the HAC writhing reaction, dispersible tablet can decrease the writhing times obviously and delay the start time of writhing.In this study, CC^-Supercritical Fluid Extraction (CO2-SFE) technique was applicated to extract, refine and reserve the effective locus and effective componentsfrom the herb medcine at the maximum scale. Regarding the extract characteristic of CO2-SFE and indication of Yuanhu Zhitong anagraph, Yuanhu Zhitong dispersible tablet was prepared so that the fast-disintegriting and fast-releasing can be realized. Quality standard was established to control the process of extraction and preparation. The feasibility and advancement of the overall experiment was evaluated through dissolution test in vitro, pharmacokinetics experiment in dog and pharmacodynamics experiment in mice.Conclusion: The purity of the extract obtained by CO2-SFE technique was higher and the solid after dehydration was less, so the CO2-SFE technique was superior to traditional alcohol refluxing method in CORYDALIS YANHUSUO W. T. WANG and ANGELICA DAHURICA extraction. The preparation form of dispersible tablet is used in clinic for relieving pain . The scientific and reasonable quality standard established can assure the stability and control of preparation. Patients' compliance is improved for the low dosage.In summary, Yuanhu Zhitong dispersible tablet prepared by utilizing CO2-SFE extracts is a safe and effective analgesic, and there is a good future in development and application in future. |
